1. Field of the Invention
This invention relates to methods and apparatus for the non-invasive quantitative determination of the presence or concentration of chemical species in a multi-component material.
2. Description of the Related Art
Analysis of samples and determination of the concentration of components contained therein is a common and important process in chemistry and biology. Of particular importance is the analysis of biological fluids such as blood, urine or saliva to determine the concentrations of various components. Also of great importance is the measurement of the concentration of various chemical species embedded within a biological material.
The chemical analysis of blood, urine and other biological fluids is crucial to the diagnosis, management, treatment and cure of a wide variety of diseases including diabetes, kidney disease and heart disease. In the case of diabetes, monitoring of blood glucose levels several times per day is a necessary feature of management of the disease for many patients. In the case of people with diseases of the circulatory system, the analysis of various blood components is of importance both in diagnosis and in treatment. For example, the level of cholesterol compounds of various types in the blood of man has a strong correlation with the probability of onset of atherosclerosis. In patients with renal insufficiencies, urine analysis provides valuable information relating to kidney function. In a related application, the concentration of alcohol in blood is known to be correlated to an individual's physical response times and can provide information relating to, for example, the individuals fitness to drive a motorized vehicle.
At present analysis of biological fluid for these and other applications is commonly invasively performed, that is by removing a sample of fluid, and subjecting it to one or more chemical tests. Typically, separate tests are required for each analyte to be measured. These tests require the use of consumable supplies and reagents and are moderately expensive. Often skilled technicians are needed to remove the fluid, and to perform the chemical tests. Frequently the tests are made in centralized clinical laboratories with resulting complexity of sample tracking, and quality control. In such circumstances there are additional problems relating to the potential change in the chemical composition of the fluid between its extraction and its analysis. Furthermore the turnaround time for such measurements can be undesirably long.
Recently there have been developed a number of moderate cost devices for the measurement of analytes using portable monitors. These devices require a drop of the fluid to be analyzed to be placed on a chemically treated substrate. This substrate is then examined by the monitor and a measurement of an analyte is produced. Typical of such devices is the family of devices for self-monitoring of blood glucose concentration such as are marketed by numerous companies in the field of diabetes management. For the limited selection of analytes for which such devices are available they offer the advantage of providing analysis in a short period of time at the patient location. Furthermore, their use eliminates the requirement for the involvement of skilled technical personnel. Although these devices have acceptable accuracy they suffer from the following limitations.
In the case of blood analysis they still require removal of blood, albeit in "finger-prick" quantities. In the case of diabetic management, the removal of blood several times daily presents associated compliance problems, particularly in children, as well as problems relating to the associated physical discomfort and potential for infection. In addition these devices are not readily compatible with continuous blood glucose monitoring such as will be required for improved diabetic management in conjunction with insulin pumps or an artificial pancreas. Furthermore these devices typically measure single analytes only, or in rare instances measure a restricted set of multiple analytes using complex and more expensive chemical or fluid management techniques.
For many applications it would be desirable to be able to make real-time measurements of analytes in biological fluids. Ideally these measurements would be made non-invasively.
An as yet unrealized goal for in-vivo monitoring of biological fluids would therefore be the development of methods and apparatus for the non-invasive, real-time measurement of analytes in a cost effective manner. For in-vitro fluid analysis the ability to make rapid measurements of single or multiple analytes could decrease analysis times, thus boosting the throughput of the clinical laboratories and reducing the cost of the analyses.
In addition to the above applications, there are a number of instances in which it is desirable to measure the local concentration of chemical species in tissue either in-vivo or in-vitro. Specific examples include the monitoring of metabolic function by measurement of tissue oxygenation as described by Jobsis (Neurol. Res. 10, 7-17, 1988), or the measurement of localized changes in tissue blood perfusion such as may be indicative of hyperplastic or neoplastic tissue. Furthermore, it is widely believed that the ability to monitor certain changes in tissue chemical composition may lead to predictive tests for various types of cancer. Examples of such changes are the development of microcalcifications, specific changes in tissue chromophore types and concentrations, and specific variation in tissue hormone levels. Consequently non-invasive methods and apparatus which enabled measurements to be made of the chemical composition of tissue samples in-vivo would also be a very important development.
One approach to non-invasively determining the composition of tissue or of a biological fluid makes use of the interaction of electromagnetic radiation with the matter under examination. It is known that electromagnetic radiation having appropriate characteristics may interact with matter in two primary ways. As it passes through the material the radiation will be scattered and a portion of it will be absorbed. Different chemical species scatter and absorb to different degrees at different wavelengths. The physical composition of the medium will also effect its interaction with the radiation. A number of methods have been proposed that use optical radiation to probe tissue or fluid samples with the goal of determining the concentration of a component of the material by making use of known characteristics of the relationship between optical absorption of the medium and wavelength.
These prior attempts generally share a number of common elements. A source of optical radiation emits light which enters the medium of interest and interacts with the medium, with the result that radiation is both absorbed and scattered by the medium. Subsequently, the attenuated light is detected after it exits the medium, and its intensity as a function of wavelength is measured. The incident light is chosen so that it contains wavelengths that are partially or wholly absorbed by the chemical species for which the concentration is to be measured. Under some conditions it is possible to use the wavelength dependence of the measured intensity of the detected light to determine the absorption coefficient of the medium as a function of wavelength. It is then sometimes possible to use techniques from the fields of chemometrics and statistics to deduce the concentration of individual and multiple analytes within the medium.
Such prior art is represented, for example, by the development of the pulsed oximeter such as is described in U.S. Pat. No. 4,621,643 (New, Jr. et al., 1986). Such a device allows the determination of the percentage of oxygen saturation of the blood (i.e. the relative saturation).
It is also represented by a series of patents, typified by U.S. Pat. No. 4,223,680 (Jobsis, 1980) and U.S. Pat. No. 4,281,645 (Jobsis, 1981), in which similar optical intensity measurements are made in order to quantify relative tissue oxygenation and metabolism using the characteristic optical absorption spectra of both haemoglobin and of the cellular enzyme cytochrome a, a.sub.3.
In both of these techniques the presence of high levels of scattering in the tissue prevents the determination of absolute concentrations of these chemical species in the blood or in the tissue. It is possible, however, to obtain relative measurements of their concentrations by making use of specific properties of the absorption spectra of the analytes and taking advantage of the fact that these analytes are present in relatively high concentrations under conditions of clinical significance. Devices based on these principles are now in wide clinical use.
The prior art is also represented by a variety of techniques directed at the non-invasive measurement of blood glucose at concentrations around the normal (fasting) physiological level of 60-115 mg/dl and of numerous other chemical components of blood of physiological significance. Typical of these techniques are those embodied in Rosenthal et al. (U.S. Pat No. 5,028,787), Robinson et al. (U.S. Pat. No. 4,975,581), Barnes et al. (U.S. Pat. No. 5,070,874), Clarke (U.S. Pat. No. 5,054,487) and other related patents.
Unfortunately, in biological samples of practical interest the scattering of the medium is sufficiently strong and wavelength dependent that it is no longer a good approximation to assume that the attenuation of the light in the medium is primarily due to absorption. Nor can scattering be considered as a wavelength independent loss mechanism. For these reasons the techniques described in the prior art result in measurement of an effective attenuation coefficient rather than of an absorption coefficient for the sample. This, combined with the fact that the scattering properties of tissue vary considerably from sample to sample, preclude the use of these prior art techniques for the accurate determination of the concentrations of analytes such as blood glucose at the relatively low concentrations typically of clinical interest.
The key problem in the use of non-invasive techniques such as those above to determine the composition of tissue or of a biological fluid is that the high degree of scattering present in the sample and its intersample variability preclude the development of an accurate algorithm relating the effective attenuation coefficient in the sample to the actual absorption coefficient in the sample. Under conditions where such an algorithm can be developed, techniques developed in the field of chemometrics and statistics have been used in many instances to enable the determination of absolute concentrations of analytes in complex media.
Such techniques have been widely used for the analysis of food and agricultural products. Typically, these techniques rely on careful sample preparation to ensure that there is negligible intersample variation in scattering properties, and then make use of empirical calibration techniques to develop a robust algorithm relating attenuation of incident radiation to absorption and thus ultimately to component concentration. In the case of biological samples such careful sample preparation is not feasible. Furthermore, typical component concentrations measured by such techniques have been in the range of 1% to 50%. The concentrations of interest for biological monitoring are often orders of magnitude smaller with the result that inaccuracies introduced to the measurement as a result of scattering are greatly magnified.
It thus appears that the high scattering, low concentrations and intersample variability inherent in the application of these techniques to biological analysis preclude the direct application of the prior art to non-invasive monitoring. It is clear by analogy to the prior art, however, that accurate deduction of the concentration of a variety of species in blood and tissue could be made using statistical and chemometric techniques if only an accurate measurement could be made of the absorption spectrum of the material rather than of its attenuation spectrum.
It has been shown in the prior art that the technique of photo-acoustic spectroscopy can be used to determine the absorption coefficient of certain types of media and that this technique can sometimes be used in the presence of high levels of scattering in the medium. As described by Rosencwaig in U.S. Pat. No. 3,948,345 the photo-acoustic effect is observed when a modulated light beam is incident on a sample and is modulated at low frequencies, usually below one kHz. As a result of the periodic heating of the material by the modulated absorption of light, thermal waves are generated in the medium, usually a solid. These thermal waves cause thermal fluctuations in a surrounding medium, usually a gas, with the result that a periodic acoustic wave, with a frequency equal to that of the light modulation, is launched into that surrounding medium. This "photoacoustic" wave can be detected, for example by means of a microphone positioned within the surrounding medium. The magnitude of the acoustic signal is determined by the degree of absorption of the radiation in the sample.
Recently this technique has been applied to the analysis of solid samples of biological material by M. G. Rockley et al (Science, 210, 21 Nov., 1980, pp 918-920). This technique of conventional photoacoustic spectroscopy requires the use of a sealed sample cell in which pressure fluctuations can be detected and measured in the gas above the sample. This type of geometry clearly precludes the use of this technique in an in-vivo situation.
In an extension of the conventional photoacoustic technique, Patel and Tam have described a specialized branch of photoacoustic spectroscopy in U.S. Pat. No. 4,303,343. This technique is quite distinct from earlier photoacoustic spectroscopic applications in that the detected acoustic wave is generated in the sample itself rather than in the surrounding medium subsequent to sample heating. In this technique a short pulse of light is incident on the sample with a pulse length that is typically of the order of 1 .mu.s but may range from 10.sup.-7 to 10.sup.-4 seconds in length. The short and intense light pulse generates a rapid heating of the sample with the resultant generation of an acoustic wave of high frequency in the sample medium itself. This acoustic wave can be detected by means of a fast acoustic detector placed in contact with the medium. Important prerequisites for the application of the technique of Patel and Tam are that the sample be relatively transparent (with an absorption coefficient of less than 10.sup.-2 cm.sup.-1); that scattering in the sample not be sufficiently great to significantly perturb the intensity distribution of the optical pulse as it travels through the sample; and that the optical beam diameter be small compared with the distance travelled in the medium by an acoustic wave during the optical pulse. Unfortunately, for biological materials of diagnostic interest, the first two of these prerequisites are generally not satisfied while the third is incompatible with measurement geometries of practical utility.
Nonetheless, it remains an important goal for the treatment of a variety of diseases, in particular of diabetes, to be able to measure the concentration of various chemical species within biological fluids or within tissue non-invasively and in real time.
It is, accordingly, the object of the present invention to provide a new and improved method and apparatus using electromagnetic radiation for the detection and quantification of various chemical species within biological media.
A specific object of the invention is to provide a new and improved method and apparatus for measuring the concentration in humans and animals of chemical species such as glucose, cholesterol, alcohol, bilirubin, ketones, fatty acids, lipoproteins, urea, albumin, creatinine, white blood cells, red blood cells, haemoglobin, blood oxygen, inorganic molecules such as phosphorous or various drugs and pharmaceutical compounds in blood, urine, saliva or other body fluids.
A further object of the invention is to provide a new and improved method and apparatus for making such measurements non-invasively, quickly, easily and at reasonable cost.
A further object of the invention is to provide a new and improved method and apparatus for measuring the concentration in tissue of chemical species such as oxygenated haemoglobin, cytochrome a, a.sub.3, insulin, glucose, bilirubin, various proteins and chromophores, microcalcifications, various hormones or drugs such as hematoporphyrin derivative (Photophrin).
A further object of the invention is to provide a new and improved method and apparatus for making such measurements non-invasively, quickly, easily and at reasonable cost.
A further object of the invention is to provide a new and improved method and apparatus for measuring the concentration of a variety of chemical components in a complex mixture of fluid media simultaneously, or consecutively within a short time duration.